Increasing evidence suggests that cytomegalovirus (CMV) infection may be a risk factor for coronary artery disease (CAD). We hypothesized that CMV mediates its atherogenic effect by promoting abnormal coronary endothelial and vasomotor function. We measured endothelium-dependent and -independent function with intracoronary acetylcholine (ACH) and nitroprusside (NP), and flow reserve with adenosine in 108 patients (pts, mean age 56 years, 55% CAD)undergoing cardiac catheterization. Doppler flow velocity was measured and coronary vascular resistance (CVR) calculated. Blood samples were tested for a)cellular response (Tc+/-)to CMV using a T-cell proliferation assay and b)for antibody response(Ab+/-)by ELISA. Compared to CMV Ab- group, Ab+ pts had a trend to a lower ACH-mediated dilation (53% vs. 46% decrease in CVR,P=0.09) and reduced flow reserve with adenosine (77% vs. 71% decrease in CVR,P=0.002), but similar NP responses(P=0.98). Because we recently showed that the risk of CAD is greatest in Ab+ pts lacking a cellular response to CMV (Ab+/Tc-), we analyzed microvascular function in this subset. Compared to others, CMV Ab+/Tc- pts had reduced dilation with both ACH (52% vs. 44% decrease in CVR, P=0.03) and adenosine (75% vs. 69% decrease in CVR, P=0.007). After multivariate analysis that included conventional risk factors for atherosclerosis, CMV Ab+/Tc- infection remained an independent predictor of the magnitude of dilation with ACH (P=0.019)and with adenosine (P=0.008). CMV infection, especially in the absence of a cellular response, impairs endothelial function and flow reserve. These findings suggest that CMV-related risk of atherosclerosis is mediated by injury to the vascular endothelium and to determinants of coronary flow reserve.